HDL receives cholesterol from tissues including arteriosclerotic vascular wall. Thus HDL is involved in cholesterol efflux accumulated in cells and serves as a risk-preventing factor for various types of arteriosclerosis including coronary arteriosclerosis. The blood level of HDL is known to serve as a useful prognostic indicator for onset of arteriosclerosis.
HDL is a complex of a protein called apoprotein and a lipid component such as a phospholipid, cholesterol and a triglyceride. HDL can be classified into subfractions, ApoE-Containing HDL and ApoE-deficient HDL, based on difference in the content of apolipoprotein E (ApoE) which is one of the components of the complex. The ApoE-Containing HDL has not only a high removability of cholesterol but also an antiplatelet action and has attracted attention as a super-good lipoprotein among the HDLs. Recently, an inhibitory agent for a cholesterol ester transfer protein (CETP) involved in increasing HDL-C has been expected to serve as a lipid-disorder therapeutic agent next to statin. A CETP inhibitory agent is known to mainly increase particularly ApoE-Containing HDL among HDLs.
As a method for measuring HDL-C, for example, a method in which HDL is separated from other lipoproteins by ultra-centrifugation and then subjected to measurement of cholesterol, and a method in which HDL is electrophoretically is separated; a lipid is stained; and its intensity of emission color is measured, are known. However, either one of these methods requires a complicated operation and is unable to treat many specimens. Because of the problems, these methods are not practically used in daily work.
As a method for measuring HDL-C; there is a method in which lipoproteins other than HDL are aggregated by adding a precipitator to a specimen; the aggregates are centrifugally removed; and cholesterol in the supernatant containing only HDL separated is measured. In this method, the reactivity to an HDL subfraction is known to vary depending upon the type of precipitator to be used herein. In a method using a precipitator such as phosphotungstic acid-magnesium and dextran sulfate-magnesium and heparin-calcium, ApoE-Containing HDL is aggregated with lipoproteins such as VLDL and LDL and centrifugally removed as a precipitation fraction and thus cannot be measured as an HDL fraction. In contrast, in a method using heparin-manganese and polyethylene glycol (PEG), ApoE-Containing HDL is not aggregated and thus is measured as HDL. In any case, a method of measuring HDL by separating HDL with a precipitator is simple compared to the ultra-centrifugation method and the electrophoresis method; however, it has a problem. Since an operation of separating HDL by adding a precipitator is included, this method is not satisfactory in view of simpleness. In addition, relatively a large amount of specimen is required.
Recently, as a method of simply quantifying HDL-C, a method of quantifying HDL-C by an automatic analyzer without a pretreatment using a precipitator, is known. More specifically, a method of specifically trapping HDL-C in the presence of a clathrate compound such as cyclodextrin by using chemically modified cholesterol esterase and cholesterol oxidase (see Patent Literature 1); a method in which lipoproteins except HDL are formed into an aggregate or a complex and thereafter cholesterol in HDL is trapped by an enzymatic reaction (see Patent Literatures 2 and 3); and a method of using a surfactant specifically acting on HDL and having an HLB of 13 to 14 (see Patent Literature 4) are known.